Arizona State University
Standard Operating Procedures Manual
Biosafety Level 2 Laboratory
of
Dr. ( )
(title)
(Institute or department and building, room location)
(date developed)
(annual review/revision date)
This is a template only, and should be modified to reflect the scope and details of the work being
performed in the individual lab. It may not include every section/subsection needed, but is meant only to
be a guide for composing your SOPs. Use search and replace function for most items in parentheses
including:
-Dr. ( )
-(insert room #)
-(name of agent)
-(name of PI)
-(describe required PPE)
-(name of disinfectant)
2
Arizona State University
Biosafety Standard Operating Procedures
(Building, room number)
Table of Contents
I. Introduction to General Safety and Training for the BSL2 Laboratory
A. Required Training
B. Administrative Procedures
C. Description of Laboratory
D. General Laboratory Safety
E. General Biosafety Cabinet Safety
F. General Accident Procedures
II. Standard Operating Procedures
A. 1000 – Containment Requirements
1001 – Laboratory Entry/Exit
1002 – Specimen Transport
1003 – Work within the Laboratory
B. 2000 – Proper Use and Maintenance of Equipment
2001 – Biological Safety Cabinets
2002 – Incubators
2003 – Centrifuges
2004 – Autoclave
2005 – Emergency Equipment
2006 – Repair and Service
C. 3000 – Operational Procedures
3001 – Inventory Control System
3002 – Working inside the Biosafety Cabinet
3003 – Working outside the Biosafety Cabinet
3004 – Removal of Equipment, Viable Samples, and Autoclavable Wastes
from the Biosafety Cabinet
3005 – Internal Clean-up, Decontamination and Waste Disposal
3006 – Maintenance of Laboratory (insert room #)
3007 – Recording of Data
D. 4000 – Experimental Procedures
E. 5000 – Safety Checks and Emergency Procedures
5001 – Training and Orientation
5002 – Personal Protective Equipment
5003 – Waste Removal from lab
5004 – Management of Spills
5005 – Management of Accidental Exposures
5006 – Medical Surveillance
5007 – Emergency Phone Numbers and Procedures
5007.1 – Emergency Phone Numbers
5007.2 – General Emergency Procedures
5007.3 – Responding to Specific Emergencies
F. 6000 – Outside Operations
6001 – Receiving
6002 – Packaging of (name of agent) for Shipment to Other Institutions
6003 – Proper Labeling and Documentation of Package for Shipping
6004 – Recordkeeping for the (name of PI) Laboratory
6005 – Validation and History for SOP Manual
G. Security
III. Protocols
IV. Safety Manuals
Appendices:
A.
B.
C.
D.
E.
Material Safety Data Sheets are maintained (location)
Miscellaneous Notes and Documents
ASU Biological Hazardous Waste Management Compliance Guidelines
Shipping Labels
Floor Diagram
PURPOSE
This manual applies to all research staff, hosted visitors and guests, volunteers, building staff,
and service staff who enter the laboratory. This manual will be reviewed annually by the
Principal Investigator or Laboratory Supervisor for changes or corrections to ensure that it is
accurate.
BACKGROUND
(name of agent)
(insert info specific to your agent including research aim and techniques, health hazards,
medical surveillance if applicable, symptoms, procedure for reporting exposure to PI,
EH&S and Campus Health Service, etc.)
I. Introduction to General Safety and Training for the BSL2 Laboratory
A. Required Training
The minimum requirements for qualification to work in the (name of PI) BSL2 lab are:
1) Current Environmental Health & Safety (EH&S) trainings including Biosafety,
Lab Safety, and Fire Safety and Prevention (also Radiation Safety if applicable)
2)
3)
(List as many as you need)
Laboratory personnel shall demonstrate the following:
(List of requirements)
-
Willingness to follow established laboratory safety guidelines and these standard
operating procedures.
This document will provide the basis of training in conjunction with EH&S training.
Dr. ( ) will provide information and arrange for training at the time of an individual's
initial assignment to the lab. Dr. ( ) will arrange for refresher training at least annually
and when there are any changes in processes or procedures.
For more information concerning training, also see the ASU Biosafety Manual on the
EH&S web site.
B. Administrative Procedures
It is the responsibility of each employee to carefully consider every action taken in the
BSL2 lab and its potential impact on possible exposure or contamination, and to follow
established Standard Operating Procedures (SOPs) and protocols diligently and without variance.
All employees will read and adhere to the ASU Biosafety Manual and to the SOP Manual
for Dr. ( ) laboratory. All employees will use pertinent sections in the Biosafety in
Microbiological and Biomedical Laboratories, CDC-NIH, 5th edition as a guideline and reference.
All employees will attend the Arizona State University EH&S courses in Laboratory Safety
Training, Biological Safety Training, Respiratory Protection Training, and Fire Safety/Fire
Prevention. Records of certification will be kept on file by Human Resources and EHS.
All employees working in the lab will be will be offered (vaccination available?).
No employee will be trained to work in the lab without the express permission of Dr. ( ).
New SOPs and protocols must be approved by the PI before initiation.
Current SOPs and protocols will be reviewed and/or revised by (name of PI) or
Laboratory Manager every ( ) months.
C. Description of Laboratory
The Laboratory is located in Building _, within Arizona State University. The lab contains
(list equipment).
Floor diagrams are displayed as Appendix E at the end of this manual. The locations of the sinks,
eyewash, safety showers, fire extinguisher, exits, and telephones are included on the diagram.
D. General Laboratory Safety
Work with the agent will be performed in (insert building and room number).
1. Laboratory employees must immediately notify the laboratory manager or PI in case of an
accident, injury, illness, or overt exposure associated with laboratory activities.
2. No eating, drinking, smoking, handling contact lenses, or applying cosmetics in the lab at any
time.
3. No animals or minors (persons under the age of 18), or immuno-compromised persons will
be allowed to enter the lab at any time.
4. Food, medications, or cosmetics should not be brought into the lab for storage or later use.
Food is stored outside in areas designated specifically for that purpose.
5. No open-toed shoes or sandals are allowed in the laboratory.
6. PPE includes gloves, lab coat, and eye protection.
7. All skin defects such as cuts, abrasions, ulcers, areas of dermatitis, etc. should be covered
with an occlusive bandage.
8. Mouth pipetting is prohibited; mechanical pipetting devices are to be used at all times.
9. All procedures are to be performed carefully to minimize the creation of splashes or aerosols.
10. Follow all manufacturer’s instructions and SOPs when using any of the laboratory equipment.
11. Wash hands:
 after removing gloves, and
 before leaving the laboratory.
12. Razor blades, scalpels, and hypodermic needles (“sharps”) should be discarded into the
“sharps” container in the biosafety cabinet. DO NOT recap needles.
13. Work surfaces will be decontaminated as needed with (name of disinfectant). Follow
manufacturer instructions for contact time.
All cultures, stocks, and other regulated wastes are decontaminated by autoclaving before
disposal. NOTE: No biological agent-containing material should be allowed into any
drain connected to the sanitary sewer system (e.g., from a sink) unless the method of
inactivation has been pre-approved by the Department of Environmental Health and
Safety. Please call 480.965.1823 to obtain pre-approval.
14.
E. General Biosafety Cabinet (“Hood”) Safety
1. Reserve time in advance to have priority for use of the BSCs. (If you use sign-up sheets,
describe the process here)
2. Turn on the blower in the cabinet at least 10 minutes before placing infectious materials into
the hood.
3. Check the certification sticker and Magnehelic gauge to verify that the biosafety cabinet is
working properly.
4. Check the air flow indicator to verify that the air flow is operating properly.
6. Gloves must be worn at all times.
7. DO NOT disrupt the airflow through the hood by placing ANY item on the grills or by opening
the door to the corridor.
8. In general, the interior of the hood should be considered to be a contaminated zone, even
though every effort is made to keep the surfaces clean, as is consistent with accepted
good microbiological practice and sterile technique.
10. Clean the inside surfaces of the BSC with (name of disinfectant) after completion of work.
Follow with 70% Ethanol or water.
11. Allow the blower to run for at least 10 minutes following use.
12. The UV light is turned on between procedures (at least 5 minutes) and at the end of the day
(all night). UV lights must be turned off before work begins in the hood. Do not look
directly at UV lights as this can cause eye damage.
F. General Accident Procedures
Spills - Apply paper towels to absorb the spill, and then soak paper towels with (name of
disinfectant). For spills outside the biosafety hood, alert others in the area. Use N95 mask if
there is a possibility of harmful aerosols.
Follow all aspects of the emergency SOPs without exception.
II. Standard Operating Procedures
A. 1000 – Containment Requirements
1001 - Laboratory
1001.1 Entering the lab to begin work
A. Put on protective outerwear, (describe required PPE).
B. Gather all materials for the experiment.
1001.2 Exiting Laboratory
Before exiting the lab, be sure that all required documentation has been completed, the
hood and work area are clean, all contaminated waste materials are disposed of properly,
and stocks have been returned to the proper storage area. Wash your hands.
1002 - Specimen Transport
Transport of biological materials to another building or lab within the same building should be
done in a covered container. If the samples are infectious, use a secondary container
and label it with the contents and a contact person/phone number.
1003 - Work within Laboratory (insert room #)
During work with (name of agent), persons must be wearing (describe required PPE).
1003.1 Preparation of primary containers of agent stock solutions and manipulations of primary
containers of agents should be conducted in a biological safety cabinet.
1003.2 The user should verify inward airflow of the biological safety cabinet before initiating
work by checking the Magnehelic.
1003.3 All work should be done within the operationally effective zone of the biological safety
cabinet.
1003.4 Care should be taken to avoid contaminating medium or other cell culture supplies with
(list possible contaminants).
1003.5 Discard pipets and tips appropriately (describe your system).
1003.6 The interior of the hood should be cleaned periodically.
1003.7 When vacuum lines are used with systems containing agents, they will be protected with
in-line filters to prevent entry of agents into the lines, and will be protected by a liquid
trap containing bleach. NOTE: No biological agent-containing material should be
allowed into any drain connected to the sanitary sewer system (e.g., from a
sink) unless the method of inactivation has been pre-approved by the
Department of Environmental Health and Safety.
Please call 480.965.1823 to obtain pre-approval.
B. 2000 – Proper Use of Equipment
2001 - Biological Safety Cabinets
2001.1 – To assure sterility inside the cabinet and establish proper air flow for containment, the
blower should be turned on at least ten minutes before infectious materials are to be put
into the biosafety cabinet.
2001.2 – Biosafety cabinets must be certified prior to use. A qualified outside contractor must
certify these cabinets annually. Check the certification sticker on the front of the unit to
verify your biosafety cabinet’s condition.
2001.3 – The biosafety cabinet air flow (“Magnehelic”) gauge should be checked (reading is
equal to approximately 0.5 inches) to assure proper operation of the cabinet before
placing any materials into it. Readings indicate relative pressure drop across the HEPA
filter. Higher readings may, therefore, indicate filter clogging. Zero readings may indicate
loss of filter integrity. In either of these cases, notify the Laboratory Manager or PI.
2001.4 – NEVER place anything over the front or rear grill of a biosafety cabinet.
2001.5 – Disrupting the airflow into the front grill allows contaminated air from inside the cabinet
to blow into the lab or directly at the person sitting at the cabinet. It also allows nonsterile air from the room to blow into the biosafety cabinet over the experiments.
2001.6 – Materials should be placed in the cabinet so as not to block air flow into the rear grill.
Leave a few inches for air to flow around objects. Any disruption of the air flow in the
cabinet decreases its effectiveness.
2001.7 – Before manipulating infectious materials, make sure that you have everything you need
in the cabinet. The fewer times you pull your hands out of the cabinet, the less
disruption of the air flow.
2001.8 – Work should be performed in the center of the work surface of the cabinet whenever
possible. Work outward progressing from clean to dirty (contaminated). However,
infectious agents should not be placed directly adjacent to or directly on the intake grills.
2001.9 – After manipulating infectious agents, make sure all containers are tightly closed.
2001.10 – All waste and disposable items generated by work in the cabinet should be left
(describe where it should be stored) until properly decontaminated or contained for
transport to the autoclave.
2001.11 – After the cabinet has been emptied, wipe inner surfaces with (name of disinfectant),
followed by 70% ethanol. Do not shut down the blower. (these instructions must be
written to accommodate your lab practices).
2001.12 – The bleach in the vacuum traps must be changed after one week of use or when the
flask is half full. NOTE: No biological agent-containing material should be
allowed into any drain connected to the sanitary sewer system (e.g., from a
sink) unless the method of inactivation has been pre-approved by the
Department of Environmental Health and Safety.
Please call 480.965.1823 to obtain pre-approval.
2001.13 – The vacuum filters must be replaced if clogged or if liquid makes contact with the
filter. Used filters should be placed in the waste to be autoclaved.
(NOTE: Please see the section on Biological Safety Cabinets in the ASU Biosafety Manual for
more information.)
NOTE: Though Class IIB cabinets are hard-ducted (so that all air is removed from the room),
Class IIA cabinets recirculate about 70% of the air inside themselves and exhaust the
remainder to the lab. Any use of volatile solvents, such as absolute ethanol, should be
kept to a minimum or done elsewhere. Dangerously high levels of volatile vapors
can accumulate inside the cabinet and pose a threat of fire or explosion.
2002 – Incubators
2002.1 – Upright Incubators (these must be written specific to your lab)
A. Incubators are normally set at 37o C.
B. Temperature should be checked each day by all users.
C. Operation manuals are located (describe where these are).
D. If an alarm is sounding, check the panel for the identifying blinking light.
1. If there is no obvious reason for the alarm, contact the Lab Manager or PI.
2. The “CO2 Low” (or High) message indicates a deviation from 5% CO2. Check
the hose from the wall to the unit.
3. The “tank farm” must be checked for empty tanks once/week.
E. Decontaminate incubators at least every (insert length of time).
2002.2 – Water bath
The water bath should be monitored for water level, and filled with distilled water only.
To prevent growth of any organisms, water should be treated with (name of
disinfectant).
2003 – Centrifuges
(describe procedures)
2004 – Autoclave
(describe procedures)
(Add similar lists for each type of equipment, such as floor shakers, microplate reader, etc.)
2005 – Emergency Equipment
2005.1 – Fire Extinguisher, located (location of fire extinguisher).
A. Operation
1. Fire extinguishers should be used only if the fire is small and confined to one
small area! USE JUDGEMENT IN THIS! DO NOT CREATE A LIFETHREATENING SITUATION WHILE TRYING TO EXTINGUISH A FIRE!
2. To operate, pull the pin to release the handle.
3. Stand at a safe distance from the fire (as directed on the fire extinguisher).
4. Aim the nozzle at the base of the fire, squeeze the handle to discharge the
agent, and sweep completely left and right until a few seconds after
seeing no fire.
B. Maintenance
Fire extinguishers are inspected annually by EH&S. Check the gauge periodically
to ensure operational status. Call EH&S at 965-1823 if you have any
questions.
2005.2 – Telephones are located (describe locations).
2006 – Repair and Service
(describe lab policy for decontaminating equipment before having service done on it)
C. 3000 – Operational Procedures
3001 – Inventory Control System
(describe how your lab keeps track of stocks and/or letting responsible person know when stocks must
be replenished).
3002 – Working inside the Biosafety Cabinet
(describe lab practices, such as use of UV lights, tip cache, disinfectant use, electrical
receptacle power on or off, etc.)
3003 – Working outside the Biosafety Cabinet
Working outside the hood includes such actions as transporting samples from the hood to a
centrifuge, incubator, sonicator, or water bath.
3003.1 – Vials or tubes being transported to the centrifuge, water bath, etc. should be in a
stable rack.
3003.2 – No liquid should be allowed into any drain connected to the sanitary sewer system
(e.g., from a sink) unless the method of inactivation has been pre-approved by the
Department of Environmental Health and Safety.
Please call 480.965.1823 to obtain pre-approval.
3004 – Removal of Equipment, Viable Samples, and Autoclavable Wastes from the Biosafety Cabinet
(insert your lab practices for removal of infectious materials and proper disposal)
3005 – Internal Clean-up and Decontamination of the Biosafety Cabinet
(describe how your lab handles disinfection)
Note that the procedures for biohazardous waste handling are described in the ASU
Biological Hazardous Waste Management Compliance Guidelines.
3006 – Maintenance of Laboratory (complete this list according to lab practices)
3006.1 Stocks are stored ……..are prepared by…….. (etc)
3006.2 Clean-up consists of ……..
3006.3 Reagents are available……….
3006.4 Organic waste bottles ………..
3006.5 The vacuum traps ……..
3007 – Recording of Data
(describe lab practices)
D. 4000 – Experimental Procedures
(include major protocols for lab procedures)
E. 5000 – Safety Checks and Emergency Procedures
5001 – Training and Orientation
(describe your lab requirements)
All employees will attend the Arizona State University EH&S courses in Laboratory Safety
Training, Biological Safety Training, and Fire Safety/Fire Prevention (Respiratory
Protection Training and Radiation Safety Training as needed), and annual refreshers.
5002 – Personal Protective Equipment (PPE)
5002.1 When using a biological safety cabinet, protective clothing, including gloves and
a long-sleeved body covering (gown, laboratory coat, smock, coverall, or similar
garment) should be worn so that hands and arms are completely covered to
prevent contamination of cultures, skin and street clothing.
5002.2 Eye protection should be worn when handling infectious organisms or chemicals.
5002.3 These requirements also apply to anyone working in the area while someone
else is working at the biosafety cabinet.
5003 – Waste Removal from the Lab
(describe your lab practices, e.g., autoclave and place in red drum for EH&S pick-up)
5004 – Management of Spills
(describe your lab practices)
Apply paper towels to absorb the spill, and then soak paper towels with (name of
disinfectant). For spills outside the biosafety hood, alert others in the area. Use N95
mask if there is a possibility of harmful aerosols.
Notify the PI. The PI will contact EH&S and Campus Health if there was an exposure.
5005 – Management of Accidental Exposures
In the event of an exposure to an infectious agent or material:





Intact skin
Remove contaminated clothing.
Vigorously wash contaminated skin for 1 minute with soap and water; there is a safety
shower (location).
Inform Dr. ( ) and seek medical attention at the ASU Campus Health Service.
Broken, cut, or damaged skin or puncture wound
Remove contaminated clothing.
Vigorously wash contaminated skin for 5 minutes with soap and water; there is a safety
shower (location).






Inform (name of PI) and seek medical attention at the ASU Campus Health Service, or
call 911 for assistance. If you are contaminated with (name of agent), notify the
emergency responders of the contamination.
Eye
Immediately flush eyes for at least 15 minutes with water, using an eyewash (location);
Hold eyelids away from your eyeball and rotate your eyes so that all surfaces may be
washed thoroughly; and
Inform Dr. ( ) and seek medical attention at the ASU Campus Health Service, or call 911
for assistance. If you are contaminated with (name of agent), notify the
emergency responders of the contamination.
Ingestion or Inhalation
Inform Dr. ( ) and seek medical attention at the ASU Campus Health Service, or call 911
for assistance. If you are contaminated with (name of agent), notify the
emergency responders of the contamination.
Do not induce vomiting unless advised to do so by a health care provider.
5006 – Medical Surveillance
(describe any available vaccines or treatments for your lab’s infectious agent, as well as
procedures to follow, symptoms to monitor for, etc.)
5007 – Emergency Phone Numbers and Procedures
5007.1 Emergency Phone Numbers
Fire and Medical Emergencies
Police
Laboratory PI
Laboratory Manager
911
911
Home Phone
Office Phone
Cell Phone
Home Phone
Office Phone
Campus Health
St. Luke’s Hospital
965-3346
(480) 968-9411
ASU Police Department
965-3456
Department of Environmental Health & Safety
(Main Number)
Biosafety Officer (EHS)
965-1823
Biosafety Officer (EHS)
965-1344
965-5389
5007.2 Emergency Procedures
In responding to an emergency situation, all persons must be aware of these basic
concerns:
Emergency Equipment:
TELEPHONES are located (insert location).
FIRE EXTINGUISHER is located (insert location).
EYEWASH is located (insert location).
SAFETY SHOWER is located (insert location)
Order of Priority in an Emergency:
I. SAFETY FOR THE EMPLOYEE/STUDENT(S) IN THE LABORATORY.
Notify anyone else present in the lab.
Safety for persons in the lab must be the first consideration. The circumstances
of the emergency will determine the feasibility of carrying out site control.
II. DANGER OF OUTSIDE CONTAMINATION (Site Control):
Do not exit building wearing contaminated clothing or protective outerwear
(gown, gloves).
Leave biosafety cabinet on.
Place covers on open containers of viable agents.
5007.3 Responding to Specific Emergencies
(describe your lab’s practices)
6001 – Receiving Reagents/Cells/Viruses
(describe lab practices for items you are likely to receive)
6002 – Packaging of (name of agent) for Shipping to Other Institutions
(list authorized person for shipping hazardous materials – specialized training is required for
shipping hazardous materials including infectious agents, contact EH&S for more information)
6003 – Proper Labeling and Documentation of Package for Shipping
(list persons responsible for labeling and preparing documentation)
6004 – Recordkeeping in the (name of PI) Laboratory
(describe required records for your lab)
6005 – Validation and History for SOP Manual (optional)
I hereby certify that I have reviewed the contents of this manual and that it reflects my current
operating policy for the laboratories ( ) located in ( ).
G. 7000 - Security
(describe the building/lab practices)
I. Protocols (attach)
II. Safety Manuals
1. ASU Biosafety Manual
2. Biosafety in Microbiological and Biomedical Laboratories, CDC-NIH, 5th edition.
Dr. ( )
Professor
(Institute/Department)
Signature _____________________________ Annual Review Date ________
History of Manual’s Creation
Date Created:
Author:
Reference Source(s):
APPENDICES
A. Material Safety Data Sheets are maintained in a notebook located (describe locations).
B. Miscellaneous Notes and Documents
C. ASU Biological Hazardous Waste Management Compliance Guidelines
D. Shipping Labels
E. Floor Diagram